Patients with advanced malignant mesothelioma (MM) have limited treatment options. The standard first-line chemotherapy regimen, pemetrexed/cisplatin, improves survival compared to cisplatin alone from 9 to 12 months. In order to develop mechanism-based therapies for MM, it is important to identify the oncogenic mutations and signaling pathways that drive its development and sustain its maintenance. Although about 75% of MM carry mutations at the NF2 locus, which encodes Merlin. We have discovered that inactivation of Merlin drives the development and maintenance of MM by activating the pro-oncogenic E3 ubiquitin Cullin Ring Ligase CRL4DCAF1, which in turn inhibits the Hippo pathway tumor suppressor kinases Lats1/2. Our Preliminary Studies indicate that the CRL inhibitor MLN4924, developed by Millenium-Takeda, exhibits exhibit selective preclinical efficacy in NF2 mutant MM cells. In addition, in vitro and in vivo studies indicate that MLN4924 significantly enhances the efficacy of pemetrexed and cisplatin in NF2 mutant MM. To establish the pre-clinical and clinical activity of MLN4924, alone and in combination with chemotherapy, in NF2 mutant MM, we will pursue the following independent, but interrelated Specific Aims: 1) To test the preclinical activity of MLN4924, alone and in combination with chemotherapy, in a novel Genetically Engineered Mouse Model (GEMM) of MM. Nf2Flox/Flox; LucR mice will be injected in their pleural cavity with lentiviruses encoding Cre in combination with short hairpin (sh) RNAs targeting the mRNAs encoded by Cdkn2a or both Cdkn2a and Bap1. Tumor-bearing mice will be treated with MLN4924, pemetrexed/cisplatinum, or both and subjected to bioluminescent imaging to monitor tumor growth. Molecular studies on tumor samples will enable us to verify target inhibition and to determine the mechanism of action of MLN4924. 2) To test the preclinical efficacy of MLN4924, alone and in combination with chemotherapy, in patient-derived xenograft (PDX) models of MPM. PDX models of MMs carrying mutations at the NF2 locus, the BAP1 locus, or at both loci will be treated with MLN4924, alone and in combination with cispatin/pemetrexed, and the results will be examined as described above. The sensitivity of prospectively isolated cancer stem cells to the drugs will be examined in tumor organoid cultures. 3) To test the clinical efficacy of MLN4924 by conducting a phase I/II trial in patients with pleural or peritoneal MM. We will conduct a single institution trial comprising two cohorts: 1 a phase II trial of single agent MLN4924 in patients with NF2 mutant MM, and 2) a phase I trial combining MLN4924 with pemetrexed/cisplatin in previously untreated patients. Information gained from pre-clinical studies in the newly developed GEMM model and state-of-the-art PDX models will help to refine the analysis of patient samples from the clinical trial and, conversely, information gained from the clinical trial will motivate additional pre-clinical studies. We envisin that mechanism-based therapies, such as the one proposed here, will radically improve the outcome of patients with MM.